Experiment
Yeast as background and USP2 proteins spiked in different concentration. This Experiment is designed for DIA benchmarking of different workflow using DIA-NN.
Import data
Reading DIA data from data/report.tsv using Precursor.Translated intensities from DIA-NN.
All the precursors with Q-value < 0.01 are used for the analysis.
Creating Q-Features object and run pre processing
The pre- processing steps are:
- Filtering precursor with more than 3 samples with no missing intensities across all the samples
- Select only proteotypic peptides
- Select proteins with at least 3 peptides
Data Missing Analysis
The plot shows the completness of the experiments at precursor and summarized level. On the the x-axis peptides/proteins are ordered based on the increasing NA content.
Normalization
Raw intensities from DIA-NN are processed as follow:
- Log 2 transformation
- Normalized across all the samples using quantiles method
- Summarization at protein level using robustSummary function
PCA
DE Analysis
Using MSqRob2.
GroupA GroupB GroupC GroupD GroupE
15.13545 14.78455 14.88787 15.01882 15.01860 QC plots
Group Comparison
This volcano plot summarizes the differential expression landscape in the comparison between the two groups
Differential Expressed Proteins 
These bar plots summarizes the number of significantly upregulated/downregulated number of miRNAs based on different adjusted p-values (selected adjusted p-values are 0.001, 0.01, 0.05, and 0.1 - see facet headers) and log2 fold-change thresholds (on the x-axis) used to define the significance levels.
This volcano plot summarizes the differential expression landscape in the comparison between the two groups
Differential Expressed Proteins 
These bar plots summarizes the number of significantly upregulated/downregulated number of miRNAs based on different adjusted p-values (selected adjusted p-values are 0.001, 0.01, 0.05, and 0.1 - see facet headers) and log2 fold-change thresholds (on the x-axis) used to define the significance levels.
Session Info
R version 4.3.3 (2024-02-29 ucrt)
Platform: x86_64-w64-mingw32/x64 (64-bit)
Running under: Windows 10 x64 (build 19045)
Matrix products: default
locale:
[1] LC_COLLATE=English_Belgium.utf8 LC_CTYPE=English_Belgium.utf8
[3] LC_MONETARY=English_Belgium.utf8 LC_NUMERIC=C
[5] LC_TIME=English_Belgium.utf8
time zone: Europe/Brussels
tzcode source: internal
attached base packages:
[1] stats4 stats graphics grDevices utils datasets methods
[8] base
other attached packages:
[1] reshape2_1.4.4 pheatmap_1.0.12
[3] biomaRt_2.58.2 gridExtra_2.3
[5] ggVennDiagram_1.5.2 hrbrthemes_0.8.7
[7] factoextra_1.0.7 MSnbase_2.28.1
[9] ProtGenerics_1.34.0 mzR_2.36.0
[11] Rcpp_1.0.12 DT_0.33
[13] plotly_4.10.4 here_1.0.1
[15] magrittr_2.0.3 lubridate_1.9.3
[17] forcats_1.0.0 stringr_1.5.1
[19] purrr_1.0.2 readr_2.1.5
[21] tidyr_1.3.1 tibble_3.2.1
[23] ggplot2_3.5.0 tidyverse_2.0.0
[25] dplyr_1.1.4 visdat_0.6.0
[27] readxl_1.4.3 msqrob2_1.10.0
[29] QFeatures_1.12.0 MultiAssayExperiment_1.28.0
[31] SummarizedExperiment_1.32.0 Biobase_2.62.0
[33] GenomicRanges_1.54.1 GenomeInfoDb_1.38.8
[35] IRanges_2.36.0 S4Vectors_0.40.2
[37] BiocGenerics_0.48.1 MatrixGenerics_1.14.0
[39] matrixStats_1.3.0
loaded via a namespace (and not attached):
[1] splines_4.3.3 later_1.3.2 bitops_1.0-7
[4] filelock_1.0.3 cellranger_1.1.0 preprocessCore_1.64.0
[7] XML_3.99-0.16.1 lifecycle_1.0.4 rstatix_0.7.2
[10] doParallel_1.0.17 rprojroot_2.0.4 lattice_0.22-6
[13] MASS_7.3-60.0.1 crosstalk_1.2.1 backports_1.4.1
[16] sass_0.4.9 limma_3.58.1 rmarkdown_2.26
[19] jquerylib_0.1.4 yaml_2.3.8 httpuv_1.6.15
[22] RColorBrewer_1.1-3 MsCoreUtils_1.14.1 DBI_1.2.2
[25] minqa_1.2.6 abind_1.4-5 zlibbioc_1.48.2
[28] AnnotationFilter_1.26.0 RCurl_1.98-1.14 rappdirs_0.3.3
[31] gdtools_0.3.7 GenomeInfoDbData_1.2.11 ggrepel_0.9.5
[34] crul_1.4.2 ncdf4_1.22 codetools_0.2-20
[37] DelayedArray_0.28.0 xml2_1.3.6 tidyselect_1.2.1
[40] farver_2.1.1 httpcode_0.3.0 lme4_1.1-35.3
[43] BiocFileCache_2.10.2 jsonlite_1.8.8 iterators_1.0.14
[46] systemfonts_1.0.6 foreach_1.5.2 tools_4.3.3
[49] progress_1.2.3 glue_1.7.0 BiocBaseUtils_1.4.0
[52] Rttf2pt1_1.3.12 SparseArray_1.2.4 xfun_0.43
[55] withr_3.0.0 BiocManager_1.30.22 fastmap_1.1.1
[58] boot_1.3-30 fansi_1.0.6 digest_0.6.35
[61] timechange_0.3.0 R6_2.5.1 mime_0.12
[64] colorspace_2.1-0 RSQLite_2.3.6 utf8_1.2.4
[67] generics_0.1.3 fontLiberation_0.1.0 data.table_1.15.4
[70] prettyunits_1.2.0 httr_1.4.7 htmlwidgets_1.6.4
[73] S4Arrays_1.2.1 pkgconfig_2.0.3 gtable_0.3.4
[76] blob_1.2.4 impute_1.76.0 XVector_0.42.0
[79] htmltools_0.5.8.1 carData_3.0-5 fontBitstreamVera_0.1.1
[82] MALDIquant_1.22.2 clue_0.3-65 scales_1.3.0
[85] png_0.1-8 knitr_1.46 tzdb_0.4.0
[88] nlme_3.1-164 curl_5.2.1 nloptr_2.0.3
[91] cachem_1.0.8 parallel_4.3.3 extrafont_0.19
[94] AnnotationDbi_1.64.1 mzID_1.40.0 vsn_3.70.0
[97] pillar_1.9.0 grid_4.3.3 vctrs_0.6.5
[100] pcaMethods_1.94.0 ggpubr_0.6.0 promises_1.3.0
[103] car_3.1-2 dbplyr_2.5.0 xtable_1.8-4
[106] cluster_2.1.6 extrafontdb_1.0 evaluate_0.23
[109] cli_3.6.2 compiler_4.3.3 rlang_1.1.3
[112] crayon_1.5.2 ggsignif_0.6.4 labeling_0.4.3
[115] affy_1.80.0 plyr_1.8.9 stringi_1.8.3
[118] viridisLite_0.4.2 BiocParallel_1.36.0 munsell_0.5.1
[121] Biostrings_2.70.3 lazyeval_0.2.2 fontquiver_0.2.1
[124] Matrix_1.6-5 hms_1.1.3 bit64_4.0.5
[127] gfonts_0.2.0 KEGGREST_1.42.0 statmod_1.5.0
[130] shiny_1.8.1.1 broom_1.0.5 igraph_2.0.3
[133] memoise_2.0.1 bslib_0.7.0 affyio_1.72.0
[136] bit_4.0.5